Introduction

Many intestinal disorders are due to intestinal parasites which cannot be diagnosed by clinically. Laboratory investigation is therefore required and the staff responsible should have adequate expertise in examining faecal specimens for parasitic organisms.

 Relevant information required

The request form should always state the patients clinical symptoms and signs and whether the patient is resident in the UK, or had recent overseas travel. If the patient has had no recent history of overseas travel, examination for Cryprosporidium, Giardia and Microsporidia if immunocompromised should be considered. If overseas travel has been undertaken, it is important to note whether the patient is ill or whether a routine post-Tropical screen is requested. The Geographical location is also important as it may indicate these parasites which could be present.

Collection of samples

If a faecal sample is not properly collected and taken care of before examination, they will be of little or no value for accurate diagnosis. This is especially true if protozoa are present. Amoebic trophozoites begin to degenerate 1-2 hours after passage, as do flagellate trophozoites. Cysts will deteriorate if the faecal specimens are left standing for many hours or overnight, especially at high temperatures.

Helminth eggs and larvae are less affected by the age of the specimen than are protozoa. Nevertheless, changes may occur that could affect their identification. eg, hookworm larvae may become embryonated and larvae may hatch from the eggs risking confusion with Strongyloides larvae. Larvae themselves may disintegrate thus making their identification difficult.

To ensure that good specimens are provided for examination, it is important to note the following points.

1. A clean dry container must be used for the collection of faecal samples. Urine and water will destroy trophozoites, if present, and the presence of dirt also causes identification problems.

2. Ideally the specimen should be brought to the lab as soon as it is passed, to avoid deterioration of protozoa and alterations of the morphology of protozoa and helminths.

3. The specimen container should be clearly labelled with the patients name, date, and time of passage of the specimen.

4. An amount of stool adequate for parasite examination should be collected and a repeat sample requested if too little is supplied.

5. Diarrhoeal specimens, or those containing blood and mucus, should be examined promptly on arrival in the laboratory. The specimens may contain motile amoebic or flagellate trophozoites and may round up and thus be missed if examination is delayed. Where amoebic dysentery is suggested, the laboratory should be informed that a "hot stool" is being supplied so that it can be examined within twenty minutes of being passed.

6. With the exception of "hot stools" if specimens cannot be examined as soon as they arrive, they should be put in the refrigerator.

 Visual observation of the faecal sample

It is important to observe the macroscopic appearance of the stool as this can give a clue to the type of organisms present. Therefore the consistency; formed, unformed or liquid; the colour and the presence or absence of an exudate are reported. The presence of adult worms can also be seen in a freshly passed stool eg adult stages of Ascaris lumbricoides and Enterobius vermicularis. Proglottids of Taenia species can also be seen.

Routine procedure for the microscopic examination of faecal samples for parasites

1. Direct microscopy should be done on all unformed and liquid samples by mixing a small amount of the specimen in 0.9% sodium chloride solution. This permits detection of trophozoites of Entamoeba histolytica and Giardia lamblia. It can also provide information on the content of the stool ie the presence of leucocytes and red blood cells.

2. A formol-ether concentrate should be done on all faecal samples examined for parasites. This reveals the presence of most protozoan cysts, eggs of nematodes, cestodes and trematodes and also the larval stages of some nematodes.

3. A permanently stained direct faecal smear should be used for all bloody, liquid or semi-formed stools. The smear can reveal the presence of intestinal parasites which can be either destroyed or missed by the formol-ether concentration method eg. Dientamoeba fragilis.

4. Specimens from patients with HIV should be left in 10% formalin for hour before proceeding with parasite examination.

Principals of diagnostic methods for the identification of parasites

The principal of the successful identification of faecal parasites is based upon

1. Measurement - The use of an eyepiece graticule is of the utmost importance, especially for cyst identification.

2. Morphology - In protozoan cysts, the number of nuclei and the presence of inclusions eg glycogen mass and chromidial bar, aid the identification of protozoa. In trophozoites, the presence of red cells in amoebae are diagnostic of Entamoeba histolytica. Flagella also aid identification of protozoan trophozoites.

3. Appearance - In helminth eggs, the shape of the egg, the thickness of the shell, the colour of the ovum and the presence or absence of features such as an operculum, spine or hooklets are diagnostic pointers to the identity of the parasite.

4. Stains, both temporary and permanent also aid in identification of the parasite. The addition of iodine to formol ether concentrates highlights the internal structure of cysts and helps distinguish between vegetable matter and cysts.. Permanently stained faecal smears are useful in demonstrating the nuclear pattern of cysts.

Reporting of parsites

Ideally, the presence of all parasites should be reported, whether they be pathogens or non-pathogens. This particularly applies to the presence of cysts. However, if it is laboratory practice to report all cysts, the report should state whether they are pathogenic or non-pathogenic.

The stage of the parasite should always be reported. For the protozoa, whether cysts or trophozoites are present; the stage of larvae as in Strongyloides; and whether adult stages or eggs of helminths are present.